Mareva Foster

The susceptibility of prosthetic biomaterials to infection

BackgroundDespite the use of a sterile technique and the administration of prophylactic antibiotics during surgical procedures, mesh infection continues to complicate the use of biomaterials. The purpose of this study was to compare the susceptibility to infection of prosthetic biomaterials in a live-animal model.MethodsThe following seven prosthetic mesh biomaterials were used in this study. Expanded polytetrafluoroethylene (ePTFE) with silver/chlorhexidine (DM+), ePTFE (DM), porcine intestinal submucosa (S), polypropylene (M), ePTFE/polypropylene (X), hyaluronate/carboxymethylcellulose/polypropylene (SM), and human acellular dermal matrix (A). Lewis rats (n = 108) underwent creation of a single ventral hernia; 105 of them were repaired with a different mesh (2-cm2 piece). Twelve pieces of each mesh were inoculated at the time of hernia repair with 108 Staphylococcus aureus (n = 84). Three pieces of each mesh were placed without bacterial inoculation (n = 21). In three animals, no mesh was placed; instead, the peritoneum of the hernia defect was inoculated (n = 3). After 5 days, the animals were killed and the mesh was explanted (peritoneum for the nonmesh control). The mesh was vortex-washed and incubated in tryptic soy broth. Bacterial counts were determined using serial dilutions and spot plates and quantified in colony-forming units (CFU) per square centimeter of mesh present in the vortex wash fluid (wash count) and the soy broth (broth count). Data are presented as the mean log10, with analysis of variance (ANOVA) and Tukey’s test used to determine significance (p < 0.05).ResultsThe DM+ material had no detectable live bacteria in the wash or broth counts in 10 of 12 tested samples (p = 0.05). Of the samples that showed bacterial growth, the peritoneum control group had a lower wash count than A (p = 0.05) and the lowest broth count of all the materials except for DM+ (p = 0.05). In addition, SM had a significantly lower wash count than A (p = 0.05), with no broth count difference. In regard to wash and broth counts, DM, M, X, SM, S, and A were no different (p = NS).ConclusionsThe DM+ material was the least susceptible to infection. Impregnation with silver/chlorhexidine killed the inoculated bacteria, preventing their proliferation on the mesh surface. Other than DM+, native peritoneal tissue appears to be the least susceptible to infection. Silver/chlorhexidine appears to be an effective bactericidal agent for use with mesh biomaterials.

Inhibition of Interferon, Cytokine, and Lymphocyte Proliferative Responses in Elite Swimmers with Altitude Exposure

To determine the immunologic consequences of athletic training at altitude, blood samples were taken at rest from 10 swimmers and 8 control nontraining but altitude-exposed members of the 1996 Australian Olympic Swimming Team, near the start and completion of a 21-day training camp at 2102 m. Blood leukocyte numbers dropped in both groups (p < 0.05), with the decrease greater in the swimmers (-38% swimmers, -3% controls). Concanavalin A (ConA)-induced blastogenesis decreased in both groups (p < 0.01), but the drop was greater in the control group (-32% swimmers, -56% controls, p < 0.05). Lipopolysaccharide (LPS)-induced blastogenesis more than doubled in both groups (281% swimmers, 249% controls, p < 0.01). Increases in mitogen-induced interleukin-1beta (IL-1beta), IL-4, and interferon-gamma (IFN-gamma) production and a decrease in IL-2 levels were observed in both groups after altitude exposure (all p < 0.05). The percentage of cells expressing HLA-DR fell (-33% swimmers, -20% controls, p < 0.01), whereas those expressing CD-4 expression increased (16% swimmers only, p < 0.01). Although training at medium-level altitude alters some immunologic parameters, the training-induced changes may be secondary to those induced by altitude alone.

Effects of 2-deoxy-D-glucose administration on immune parameters in mice.

Physical exercise and diet alterations have been shown to affect immune parameters. Similar effects are also induced by the administration of the non-metabolizable glucose analog, 2-deoxy-D-glucose (2-DG). The current study was designed to characterize the effects of glucoprivation induced by 2-DG administration on leukocyte subset distribution and function. BDF1 mice (n = 8 per group) were injected intraperitoneally one or three times with 0, 500, 750, 1000 or 1500 mg/kg of 2-DG. Two hours after the last injection of 2-DG, immunological parameters were analyzed. A dose-dependent increase in plasma glucose concentrations of mice injected once with up to 1500 mg/kg of 2-DG was observed (p < 0.001). After either one or three injections of up to 1500 mg/kg of 2-DG, corticosterone levels, leukocyte counts in the spleen, and CD3+ cells in the thymus increased. In vitro proliferation of partially purified lymphocytes from the spleen in the presence of both concanavalin-A and lipopolysaccharide decreased in a dose dependent manner (p < 0.05). In addition, after three injections, the proportion of both thymocytes and splenocytes bearing alphabeta-TCR increased as the concentration of 2-DG increased (p < 0.01). These results demonstrate that 2-DG administration induced dose-dependent changes in both thymus and spleen cell distribution and function.

Angiogenic and immune parameters during recombinant interferon-α2b adjuvant treatment in patients with melanoma

As an adjuvant therapy for patients with high risk of recurrent melanoma, high-dose interferon (IFN)-alpha2b therapy has been shown to have some efficacy. We examined 22 patients with resected melanoma who were treated with repeated injections of recombinant IFN-alpha2b during the treatment. Both angiogenic and immune parameters were measured. White blood cells (WBCs) and lymphocyte numbers, lymphocyte subpopulations, serum concentrations of IFN-alpha and anti-IFN-alpha antibodies, and the serum vascular endothelial growth factor (VEGF), interleukin (IL)-8, and basis fibroblast growth factor (bFGF) concentrations were determined over time in resected, recurrence-free patients with American Joint Committee on Cancer (AJCC) stage III melanoma with one or less (LN+ < or = 1, n = 7) or more than one (LN+ > 1, n = 8) lymph nodes involved, and AJCC stage IV resected disease (n = 7). Follow-up and recurrence-free intervals were longer in stage III (LN+ < or = 1) patients compared with stage IV patients (P < 0.05). The number of WBCs and lymphocytes decreased during the treatment for all patient groups (P < 0.001). In addition, percentages of CD8 and CD20 were higher in stage IV patients than in stage III (LN+ > 1) and stage III (LN+ < or = 1) patients at the beginning of therapy (P < 0.05). A significant increase in the percentage of CD20+ cells, mostly B lymphocytes, was observed in the stage III (LN+ > 1) and stage III (LN+ < or = 1) patients over time but not in stage IV patients (P < 0.001). Low IL-8 and bFGF concentrations at the beginning of therapy were associated with significantly longer recurrence-free survival (P < 0.05). These results warrant a larger trial to determine if the differences observed in patients before treatment can provide prognostic markers in patients receiving IFN-alpha2b therapy.

Immune Alterations in Male and Female Mice after 2-Deoxy-d-Glucose Administration

Administration of 2-deoxy-D-glucose (2-DG) induces acute cellular glucoprivation. In the current study, we examined differences in immune parameters after 2-DG administration in both sexes. Male and female BDF1 mice were injected three times, 48 h apart, either with a saline solution (control group) or with 2-DG in saline (500 mg/kg). Two hours after the last injection, blood and spleens were collected. Plasma levels of interleukin-1beta, and interferon-gamma levels were measured. Additionally, the levels of the specific leukocyte antigens CD3, CD4, CD8, T cell receptor (TCR) alpha/beta, I-Ad, and H-2Ld/H-2Db were evaluated by flow cytometry on both blood and spleen cells. The blastogenic response of leukocytes from both tissues to mitogens was assessed. Levels of glucose, corticosterone, testosterone, progesterone, 17beta-estradiol, follicle-stimulating hormone, and luteinizing hormone were also determined. Increases in the percentage of cells bearing TCR alpha/beta and I-Ad in the blood and H-2Ld/H-2Db in the spleen were observed in the 2-DG-treated group for both sexes. In contrast, higher corticosterone and IL-1beta plasma concentrations, as well as higher percentages of splenocytes bearing TCR alpha/beta and I-Ad, and lower mitogen-induced proliferation of mature T splenocytes (79%) were observed in female but not in male mice injected with 2-DG compared with those injected with saline (p < 0.05). Taken together, these results suggest that female mice are more sensitive than male mice to immune alterations induced by 2-DG administration.

Inhibitory effects of fusarochromanone on melanoma growth

Fusarochromanone is a toxic metabolite produced by Fusarium equiseti, a fungus present in decaying cereal plants in northern latitudes; it has been detected in various food grains. Fusarochromanone has been shown to have both stimulatory and inhibitory effects on various mammalian cells, depending on the concentration used. Whether these cytotoxic effects can be used in the clinical treatment of tumors remains to be established. Here, we evaluated the effects of fusarochromanone on the growth of human melanoma cells both in vitro and in vivo. In vitro, low concentrations (0.1–1 nmol/l) of fusarochromanone were found to be cytotoxic to many melanoma cell lines. In contrast, growth of normal melanocytes was inhibited only at much higher fusarochromanone concentrations (100–200 nmol/l). In vivo, the growth of melanoma cells implanted subcutaneously in immuno-compromised mice was significantly (P<0.05) reduced by daily administration of fusarochromanone. Immunohistological analyses indicated a significant (P<0.05) increase in the expression of active caspase-3 in tumor masses of mice treated with fusarochromanone, compared with controls. Together, these observations show that fusarochromanone increased apoptosis of tumor cells and reduced tumor growth in vivo. Therefore, the effects of fusarochromanone warrant further investigation as an adjuvant molecule to prevent growth and recurrence of melanomas.

Effects of 2-deoxy-D-glucose administration on cytokine production in BDF1 mice.

Physical exercise and diet changes have been shown to affect immune parameters, and similar effects are also induced by the administration of a nonmetabolizable glucose analog, 2-deoxy-D-glucose (2-DG). The present study was designed to characterize the effects of glucoprivation induced by 2-DG administration on concentrations of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-6 in the blood and interferon-gamma (IFN-gamma), IL-2, and IL-4 in vitro production by partially purified T splenocytes in BDF1 mice. Mice (n = 8 per group) were injected intraperitoneally one or three times with 0, 500, 750, or 1000 mg/kg of 2-DG, and blood and spleens were collected 2 h after the last injection. Partially purified T splenocytes were cultured 24 h in the presence of concanavalin A (ConA). A significant increase in the corticosterone levels with the amount of 2-DG injected was observed after one or three injections (p<0.05). The amount of 2-DG injected was associated with an increase in TNF-alpha, IL-1beta, and IL-6 concentrations in the blood of mice after one or three injections of 2-DG (p<0.05). A significant decrease in in vitro proliferation of partially purified splenocytes in the presence of ConA was associated with a decrease in IFN-gamma production in the culture supernatants and an increase in IL-1 receptor expression on the cell surface (p<0.05).

Immune alterations in three mouse strains following 2-deoxy-d-glucose administration

Using 2-deoxy-D-glucose (2-DG)-induced stress, our laboratory has developed studies to define stress effects on immune responses. Here, we report effects of increasing doses of 2-DG on the immune response of BALB/c, C57BL/6 and BDF(1) mice 2 h after three injections of 0 to 2000 mg/kg of 2-DG. Female 4- to 5-week-old mice were euthanized and blood and spleens were collected. A suspension of partially purified mature T splenocytes was obtained by negative selection using J11.d2 antibodies. Glucose and corticosterone levels were measured in the plasma of each mouse. Splenocyte and mature T splenocyte suspensions were tested in in vitro proliferation assays with or without concanavalin A. Splenocytes were analyzed for the following cell-surface markers: CD3, TCR alpha/beta, CD4, CD8 and major histocompatibility complex (MHC) Class II. Significant increases in blood glucose levels were observed in C57BL/6 and BALB/c strains with the highest 2-DG dose (p<0.05). Corticosterone levels were higher in BDF(1) mice and C57BL/6 mice following the administration of 1000 and 2000 mg/kg of 2-DG, respectively (p<0.01). In vitro proliferation of mature T splenocytes in the presence of concanavalin A was decreased in BDF(1) (p<0.05) but not in BALB/c and C57BL/6 mice. In addition, in BDF(1) mice the decrease was highly correlated with an increase of CD3+ and TCR alpha/beta+ cells in the spleen. These results demonstrated high variability in the response of different mouse strains to 2-DG-induced stress.